消散石英晶体微天平对木瓜蛋白酶水解唾液膜的动力学研究

目的：本实验以人全唾作为唾液获得性膜的体外实验模型,研究不同pH值、离子强度和温度条件下木瓜蛋白酶水解的动力学。方法：通过消散石英晶体微天平在各种条件下观察酶水解唾液膜的频率变化。结果：在各种条件下唾液膜被木瓜蛋白酶部分水解。木瓜蛋白酶与唾液蛋白质的相互作用对pH值、离子强度和温度的改变敏感。在选择和应用酶时,该特性应加以考虑。降低pH值,提高离子强度（1～5mmol／L范围内）和温度,木瓜蛋白酶的催化效率增强。结论：酶动力学显示木瓜蛋白酶水解唾液膜的过程具有pH值、离子强度和温度的依赖性特点。在牙膏、漱口水、义齿清洁剂的配方中利用木瓜蛋白酶的这一特点,有助于获得理想的抗菌复合剂。酶的最佳化尚需更进一步理解反应过程中的调节机制和多重的影响因素。     

木瓜蛋白酶水解对β-酪蛋白膜的形态和黏弹性的影响

目的：评价木瓜蛋白酶水解对牛β-酪蛋白（β-CN）膜表面形态和黏弹性的影响。方法：将β-CN通过自组装固定于芯片表面,采用消散因子石英晶体微天平（quartz crystal microbalance with dissipation,QCM-D）测量β-CN膜能量消散因子（△D）和频率（△F）和黏弹性变化。β-CN膜水解前后的表面形态变化通过原子力显微镜（atomic force microscopy,AFM）表征。结果：伴随着酶浓度提高,残余的β-CN膜的黏弹性增加（P＜0．05）。AFM图像显示水解后残余的β-CN膜的厚度和粗糙度均降低。结论：木瓜蛋白酶水解β-酪蛋白β-CN膜的反应具有浓度依赖性特征;水解反应使膜变得较薄而光滑,且更加紧凑和较硬。     

不同条件下木瓜蛋白酶对唾液／色素复合膜的清除效果

目的：考察3种条件（离子强度、pH值和温度）下木瓜蛋白酶对羟基磷灰石（hydroxyapaptite,HA）表面人全唾液（whole saliva,WS）／茶黄素（theaflavin,TF）复合膜的水解效率。方法：通过消散石英晶体微天平（quartz crystal microbalance with dissipation,QcM—D）在芯片HA的表面构筑ws／TF复合膜,运用玻尔兹曼方程评估不同离子强度、pH值和温度下酶水解ws／TF复合膜的水解参数。结果：木瓜蛋白酶水解复合膜效率的离子强度依次为10mmol／L〉100mmol／L〉70mmol／L〉50mmol／L〉30mmol／L（P〈0．05）;pH值依次为pH7．O〉pH6．5〉pH7．5（P〈0．05）;温度依次为35℃〉25℃〉20oC（P〈0．05）。结论：木瓜蛋白酶水解HA表面wS／TF复合膜的效率具有离子强度、pH值和温度依赖性。     

木瓜蛋白酶水解对β-酪蛋白膜的形态和黏弹性的影响

将β-CN通过自组装固定于芯片表面,采用消散因子石英晶体微天平(quartz crystal microbalance with dissipation, QCM-D)测量β-CN膜能量消散因子(ΔD)和频率(ΔF)和黏弹性变化。β-CN膜水解前后的表面形态变化通过原子力显微镜(atomic force microscopy,AFM)表征。结果:伴随着酶浓度提高,残余的β-CN膜的黏弹性增加(P<0.05)。AFM图像显示水解后残余的β-CN膜的厚度和粗糙度均降低。结     

3种巯基蛋白酶对去磷酸化牛乳β-酪蛋白和茶黄素复合膜的清除

目的：采用茶黄素（theaflavin,TF）和去磷酸化牛p一酪蛋白（dephosphorylated bovine β—casein,Dβ—CN）在芯片表面形成蛋白质／色素复合膜的模型,评估3种巯基蛋白酶（cysteine proteases,CPs）,木瓜蛋白酶、菠萝蛋白酶和无花果蛋白酶,对DB—CN／TF复合膜的清除作用。方法：在消散石英晶体微天平（quartz crystal mi—crobalance with dissipation,QcM—D）上建立D8一CN／TF复合膜,Boltzman方程评估3种CPs水解复合膜的效率,水解前后膜变化通过原子力显微镜（atomic forc emicroscopy,AFM）和接触角表征。结果：3种CPs水解复合膜的效率由大到小依次为无花果蛋白酶、木瓜蛋白酶、菠萝蛋白酶（P〈0．05）。AFM图和接触角表征结果证实,CPs能够有效地清除结合在DG—CN表面的TF。结论：CPs具有在牙色渍清除中潜在的应用价值。在开发口腔保健产品中该类酶值得进一步研究。     

木瓜蛋白酶水解β-酪蛋白膜的动力学研究

目的：采用消散因子石英晶体微天平（QCM-D）原位、实时和动态监测木瓜蛋白酶水解β-酪蛋白生物膜的动力学过程,比较不同浓度酶的水解效率。方法：用β-酪蛋白在QCM-D上建立体外蛋白膜模型,采用不同浓度的木瓜蛋白酶水解膜,观察频率和厚度的改变,运用Boltzmann方程对实验数据进行拟合,单因素方差分析和SNK-q检验比较水解参数B、Δh、V50和C。结果：随着酶浓度增大,膜的厚度Δh减薄,ΔF上升增多,水解参数B增大,C减小,V50缩短（P〈0.05）。结论：在一定浓度范围内水解蛋白生物膜时,酶的水解效率随浓度增大而提高。     

消散因子石英晶体微天平监测人唾液在金表面形成生物膜的过程

目的:采用消散因子石英晶体微天平(QCM-D)原位、实时和动态测量全唾液、腮腺唾液和颌下腺/舌下腺唾液在Au石英晶体芯片表面形成生物膜的过程.方法:收集刺激性的上述3种唾液,QCM-D监测唾液蛋白吸附于Au石英晶体芯片表面所造成的频率改变和消散因子的变化,及形成生物膜的质量、厚度、剪切弹性模量和剪切粘度.数据用单因素方差分析和LSD检验进行两两比较(a=0.05).结果:全唾液、腮腺唾液和颌下腺/舌下腺唾液蛋白质吸附达饱和状态时的频率改变,生物膜厚度和质量为SMSLS>WS>PS(P<0.05).消散因子的变化、生物膜的剪切弹性模量和剪切粘度为PS>SMSLS和WS(P<0.05).结论:3种唾液吸附动力学特性和形成生物膜的差异是由于其成分不同所造成的.消散因子石英晶体微天平是研究膜的有效工具.     

绿茶多酚导致口腔收敛性感觉的吸附动力学研究

目的:利用吸附动力学原理和模型,探索绿茶多酚的典型化合物表没食子儿茶素没食子酸酯(EGCG)引起口腔收敛性感觉的分子机制.方法:通过消散因子石英晶体微天平测量EGCG吸附于全唾液(WS)和腮腺唾液(PS)表面的质量、厚度、弹性和黏度,考察EGCG单层膜的特性;计算Langmuir 和Fruendlich 模型的吸附常数...     

pH值、温度和离子强度对色素与人类唾液相互作用的影响

目的:通过表面等离子体共振(surface plasma resonance,SPR)技术在分子水平上原位、实时和动态考察pH值、温度和离子强度对茶黄素(theaflavin,TF)、姜黄素(curcumin,Cur)和矢车菊素(cyanidin,Cy)吸附于人全唾液(human whole saliva,WS)表面的影响。方法:利用表面等离子共振技术和吸附动力学原理,在SPR芯片表面自组装形成人全唾液蛋白质生物膜。通过改变实验条件,在不同的pH值、温度和离子强度条件下监测TF、Cur和Cy吸附于WS表面的响应强度。统计采用双因素方差分析和SNK-q检验,检验水平α=0.05。结果:伴随着pH值和温度的升高,三种色素在人类全唾液表面的响应强度降低,吸附量逐渐减少。当反应体系的离子强度位于5.0～12.5 mmol/L范围内时,吸附量与离子强度呈正相关,而当反应体系的离子强度低于5.0 mmol/L或高于12.5 mmol/L时,吸附量与离子强度呈负相关。结论:静电反应是色素和唾液蛋白质分子之间相互作用的主要驱动力。     

人工唾液条件下3种自酸蚀粘接剂与牙本质粘接耐久性研究

目的　探讨Single bond™ Universal等3种不同类型自酸蚀粘接剂与牙本质的粘接强度和耐久性。方法　选取 2016年5—7月解放军总医院口腔颌面外科拔除的24颗新鲜人离体第三磨牙,随机分为3组,每组8颗,分别选用Clearfil SE Bond（SE组）、G-Bond plus（GBp组）、Single bond™ Universal（SBU组）等3种自酸蚀粘接剂与新鲜牙本质粘接并制备试件。每组挑选出完整试件160个,再分为两个亚组（各80个试件）,分别在即刻与人工唾液浸泡6个月后行微拉伸粘接强度实验,体视显微镜下观察试件断裂类型。 结果　SE组、GBp组、SBU组即刻牙本质粘接强度分别为（55.70 ± 11.07）、（52.85 ± 7.54）和（62.82 ± 8.70）MPa,其中SBU组高于SE组和GBp组,差异有统计学意义（P < 0.05）。人工唾液保存6个月后,SE组、GBp组、SBU组牙本质粘接强度分别为（13.51 ± 4.99）、（43.53 ± 6.82）和（49.13 ± 6.58）MPa,3组之间差异均有统计学意义（P < 0.05）。6个月时粘接界面观察,SE组断裂模式均为粘接界面断裂;GBp组16个试件为牙本质内聚断裂,4个试件为复合树脂内聚断裂;SBU组28个试件为牙本质内聚断裂,8个试件为复合树脂内聚断裂。 结论　Single bond™ Universal即刻牙本质粘接强度和人工唾液老化后6个月粘接强度均显著高于Clearfil SE Bond、G-Bond plus;Single bond™ Universal粘接强度大,稳定耐久,可提供良好的粘接效果。     

Effect of various pH values, ionic strength, and temperature on papain hydrolysis of salivary film

Stimulated human whole saliva (WS) was used to study the dynamics of papain hydrolysis at defined pH, ionic strength, and temperature with the view of reducing an acquired pellicle. A quartz crystal microbalance with dissipation (QCM-D) was used to monitor the changes in frequency caused by enzyme hydrolysis of WS films, and the hydrolytic parameters were calculated using an empirical model. The morphological and conformational changes of the salivary films before and after enzymatic hydrolysis were characterized by atomic force microscopy (AFM) imaging and grazing-angle Fourier transform infrared (GA-FTIR ) spectra, respectively. The characteristics of papain hydrolysis of WS films were pH-, ionic strength-, and temperature-dependent. The WS films were partially removed by the action of papain, resulting in thinner and smoother surfaces. The infrared data suggested that hydrolysis-induced deformation did not occur on the remnants of salivary films. The processes of papain hydrolysis of WS films can be controlled by properly regulating pH, ionic strength, and temperature.     

胃食管反流病患者唾液腺功能的变化

目的探讨胃食管反流病（gastroesophageal reflux disease,GERD）患者的非刺激性唾液（unstimulated whole saliva,UWS）和刺激性唾液（stimulated whole saliva,SWS）初始pH值、唾液缓冲能力和唾液流率等唾液腺功能的改变。方法收集57例确诊的GERD住院患者和24名健康人的非刺激性唾液、刺激性唾液各2ml,测定唾液的pH值、唾液缓冲能力和唾液流率。结果①GERD组刺激性唾液的初始pH值极显著高于非刺激性唾液（P〈0.001）;②对照组非刺激性唾液的缓冲能力显著高于GERD组非刺激性唾液的缓冲能力（P〈0.01）,对照组刺激性唾液的缓冲能力高于GERD组刺激性唾液的缓冲能力（P〈0.05）;③对照组非刺激性唾液的流率极显著高于GERD组非刺激性唾液的流率（P〈0.001）,对照组刺激性唾液的流率显著高于GERD组刺激性唾液的流率（P〈0.01）。结论GERD患者唾液腺功能降低。     

咀嚼麦芽糖醇口香糖对唾液流率及pH值的影响

目的 探讨咀嚼麦芽糖醇口香糖对唾液流率与pH值的影响.方法 选10例志愿者,分别检测咀嚼麦芽糖醇口香糖或木糖醇口香糖前后不同时间段唾液的流率和pH值.应用SPSS 12.0软件分析.结果 咀嚼2种口香糖10 min内,唾液流率显著增加,在2～4 min时达到峰值,实验组和对照组分别为(4.24±0.30)mg/min和(4.53±0.19)mg/min.唾液pH值在2～4 min时上升至(7.16±0.15)和(7.02±0.20).两组的唾液流率与pH值之间差异均无统计学意义(P>0.05).结论 咀嚼麦芽糖醇口香糖也能促进唾液分泌,提高唾液pH值.     

Relationship of the unstimulated whole saliva flow rate and salivary gland size estimated by magnetic resonance image in healthy young humans

OBJECTIVE: The aim of this study was to examine the relationships between gland sizes and the flow rate and composition of the unstimulated whole saliva in humans. DESIGN: In 28 healthy young adults, the sizes of the three major salivary glands were estimated by use of a magnetic resonance (MR) imaging technique. Unstimulated whole saliva was collected for 5 min by the spitting method, and the flow rate and the concentrations of total protein, Na(+) and K(+) and pH were measured. RESULTS: The estimated sizes of the parotid and submandibular glands showed a significant positive correlation with the flow rate and the secretion rate of total protein in the unstimulated whole saliva, but that of the sublingual glands did not. Concerning the concentrations of Na(+) and K(+) and pH, there were no correlations with the salivary gland sizes. CONCLUSIONS: The results suggest that the larger the sizes of the parotid and submandibular glands, the faster the fluid flow and protein secretion rates in unstimulated whole saliva.     

碱性pH对浮游状态和生物膜状态粪肠球菌活性的影响

目的:比较生物膜状态与浮游状态下粪肠球菌对碱的耐受性。方法:制备浮游状态和生物膜状态的粪肠球菌细胞,用pH值分别为7、8、9、10、11和12的培养液作用2h,利用MTT比色法比较2种状态下细菌细胞活性变化。采用SAS6.12软件包对数据进行统计学分析。结果:低碱性环境(pH值7～9)对粪肠球菌的生长无明显影响,高碱性环境(pH值>10)下存活细菌的比例明显减少;高pH环境下,生物膜状态的粪肠球菌存活细菌比例显著高于浮游状态下细菌的存活比例。结论:粪肠球菌对碱性环境具有强耐受性,形成生物膜是粪肠球菌抵抗高碱性环境的一个重要原因。     

The effect of saliva specimen collection, handling and storage protocols on hepatitis C virus (HCV) RNA detection by PCR

OBJECTIVES: Commercial assays can now be adapted to detect salivary anti-hepatitis C virus (HCV) antibodies, increasing the potential of saliva as a non-invasive diagnostic specimen suited to surveillance and epidemiological studies. However, current diagnostic algorithms involve confirmation of HCV infection by RT-PCR. Manipulation and storage conditions of serum can influence the stability of viral RNA. This study examined whether varying specimen collection, handling and storage protocols also affected subsequent HCV RNA detection by RT-PCR applied to saliva specimens. METHODS: Whole unstimulated saliva, together with saliva samples collected in two commercially available devices (Salivette and Omnisal) were obtained from 50 HCV seropositive intravenous drug users. The specimens were subjected to a number of handling and storage conditions, including heat treatment and prolonged storage, then examined for HCV RNA by RT-PCR using primers derived from the 5' non-coding region (5'NCR). RESULTS: HCV RNA was detected in saliva samples from 25 (50%) of the patients. No single collection device or handling procedure identified all the subjects with HCV RNA positive saliva though whole saliva yielded the greatest number of positive results. CONCLUSIONS: Collection and processing of saliva specimens for RT-PCR analysis is complex. At present, detection of salivary HCV RNA by PCR is not sufficiently sensitive for use as a diagnostic tool in epidemiological studies.     

Effects of pH on the glucose and lactate metabolisms by the washed cells of Actinomyces naeslundii under anaerobic and aerobic conditions

Effects of pH on the glucose and lactate metabolism by the washed cells of Actinomyces naeslundii genospecies 1 and 2 under anaerobic and aerobic conditions were studied. The rate of acid production from glucose was the highest at pH 7.0 and decreased as the pH lowered to 4.5, irrespective of atmospheric conditions. The anaerobic end-product in the absence of bicarbonate was mainly lactate, while in the presence of bicarbonate the rate of acid production increased 1.8-2.5 times with the production of formate, acetate and succinate in addition to lactate. Under aerobic conditions, the cells produced acids from glucose along with oxygen consumption and the end-product was mainly acetate. In contrast to the glucose metabilism, the cells produced base from lactate along with oxygen consumption. The rates of base production and oxygen consumption were the highest at pH 5.5. The end-products from lactate were acetate and pyruvate. These results indicate that oral actinomyces has a various activity of glucose and lactate metabolism at a wide range of environmental pH and suggest its flexibility in surviving in dental plaque, where the environmental factors fluctuate.